Development and validation of a flax (Linum usitatissimum L.) gene expression oligo microarray.
Development and validation of a flax (Linum usitatissimum L.) gene expression oligo microarray.
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DOI：
10.1186/1471-2164-11-592
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发表时间：
2010-10-21
期刊：
影响因子：
4.4
通讯作者：
Thomasset B
中科院分区：
文献类型：
--
作者：
Fenart S;Ndong YP;Duarte J;Rivière N;Wilmer J;van Wuytswinkel O;Lucau A;Cariou E;Neutelings G;Gutierrez L;Chabbert B;Guillot X;Tavernier R;Hawkins S;Thomasset B
亚麻（Linum usitatissimum L.）已经被种植了约9000年，因此是最古老的栽培物种之一。如今，亚麻仍然因其油（油用亚麻或亚麻籽品种）以及富含纤维素的纤维（纤维用亚麻品种）而被种植，这些纤维用于高价值的亚麻衣物和复合材料。尽管亚麻衍生产品在工业上有广泛应用，而且我们对木纤维生产和油脂生物合成的调控有了实际了解，但在这两个领域还必须获取更多信息。基因组学的最新进展现在为提高我们对这些复杂过程的基础知识提供了机会。在本文中，我们报道了一种用于亚麻基因表达分析的高密度寡核苷酸微阵列平台的开发和验证。
从亚麻内部茎、外部茎、种子、叶片和根部获取的9种不同的RNA样本被用于通过大规模平行焦磷酸测序产生1066481个表达序列标签（EST）的集合。这些序列被组装成59626个单基因，并且选择了48021个序列用于寡核苷酸设计以及高密度微阵列（Nimblegen 385K）的制作，每个单基因有8个不重叠的25聚体寡核苷酸。18个独立实验被用于评估杂交质量、精密度、特异性和准确性，所有结果都证实了我们微阵列平台的高技术质量。使用定量实时逆转录聚合酶链反应（qRT - PCR）对微阵列数据进行交叉验证。根据微阵列结果选择了9个靶基因，它们反映了倍数变化的整个范围（不同样本中上调和下调的基因）。在qRT - PCR和微阵列结果中，比较所有生物学重复中每个靶基因的表达水平，获得了具有统计学意义的正相关。进一步的实验说明了我们的阵列检测各种亚麻组织以及两个对比鲜明的亚麻品种之间差异基因表达的能力。
所有结果表明，我们的高密度亚麻寡核苷酸微阵列平台可以用作一种非常灵敏的工具，用于分析多种组织以及不同品种中的基因表达。此外，这个高度可靠的平台还可用于不同亚麻组织中mRNA转录谱的定量。
Flax (Linum usitatissimum L.) has been cultivated for around 9,000 years and is therefore one of the oldest cultivated species. Today, flax is still grown for its oil (oil-flax or linseed cultivars) and its cellulose-rich fibres (fibre-flax cultivars) used for high-value linen garments and composite materials. Despite the wide industrial use of flax-derived products, and our actual understanding of the regulation of both wood fibre production and oil biosynthesis more information must be acquired in both domains. Recent advances in genomics are now providing opportunities to improve our fundamental knowledge of these complex processes. In this paper we report the development and validation of a high-density oligo microarray platform dedicated to gene expression analyses in flax. Nine different RNA samples obtained from flax inner- and outer-stems, seeds, leaves and roots were used to generate a collection of 1,066,481 ESTs by massive parallel pyrosequencing. Sequences were assembled into 59,626 unigenes and 48,021 sequences were selected for oligo design and high-density microarray (Nimblegen 385K) fabrication with eight, non-overlapping 25-mers oligos per unigene. 18 independent experiments were used to evaluate the hybridization quality, precision, specificity and accuracy and all results confirmed the high technical quality of our microarray platform. Cross-validation of microarray data was carried out using quantitative qRT-PCR. Nine target genes were selected on the basis of microarray results and reflected the whole range of fold change (both up-regulated and down-regulated genes in different samples). A statistically significant positive correlation was obtained comparing expression levels for each target gene across all biological replicates both in qRT-PCR and microarray results. Further experiments illustrated the capacity of our arrays to detect differential gene expression in a variety of flax tissues as well as between two contrasted flax varieties. All results suggest that our high-density flax oligo-microarray platform can be used as a very sensitive tool for analyzing gene expression in a large variety of tissues as well as in different cultivars. Moreover, this highly reliable platform can also be used for the quantification of mRNA transcriptional profiling in different flax tissues.
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影响因子：
6.9
作者：
Gutierrez, Laurent;Conejero, Genevieve;Van Wuytswinkel, Olivier
通讯作者：
Van Wuytswinkel, Olivier
影响因子：
5.3
作者：
Hotte NS;Deyholos MK
通讯作者：
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影响因子：
3
作者：
De Pauw, Mary A.;Vidmar, John J.;Deyholos, Michael K.
通讯作者：
Deyholos, Michael K.
影响因子：
3.3
作者：
Farmer, C.;Petit, H. V.;Capuco, A. V.
通讯作者：
Capuco, A. V.
影响因子：
4.3
作者：
Day, A;Ruel, K;Chabbert, B
通讯作者：
Chabbert, B